New inhibitors of calpain prevent degradation of cytoskeletal and myelin proteins in spinal cord in vitro.
We have determined the effects of the calpain inhibitors AK275 and AK295 upon purified m-calpain and calcium-mediated degradation of neurofilament protein (NFP) in rat spinal cord in vitro. After incubation, the soluble radioactivity and/or extent of myelin basic protein (MBP) or NFP degradation was determined. Fifty percent of caseinolytic activity was inhibited by both inhibitors at 0.6 microM concentration, while more than 90% inhibition was seen at 1.6 microM. In contrast, 37% and 64% inhibition of MBP degradation was seen with AK295 and AK275, respectively, at 10 microM concentration. The extent of NFP degradation in spinal cord was quantified from immunoblot enhanced chemiluminescence. The calcium-mediated breakdown of NFP was inhibited by both AK275 and AK295, and the inhibition was dose-dependent. A 50% inhibition of NFP degradation was seen with AK295 at 10 microM and was almost completely inhibited at 25-50 microM. AK295 was slightly more potent than AK275. These studies suggest that these potent calpain inhibitors may be used therapeutically to provide neuroprotection in vivo in experimental central nervous system trauma and ischemia.
James
T
T
Department of Neurology, Medical University of South Carolina, Charleston 29425, USA.
Matzelle
D
D
Bartus
R
R
Hogan
E L
EL
Banik
N L
NL
D000818:Animals; D002154:Calpain / Q000037:antagonists & inhibitors*; D002364:Caseins / Q000378:metabolism; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D004151:Dipeptides / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006207:Half-Life; D009185:Myelin Proteins / Q000378:metabolism*; D018696:Neuroprotective Agents / Q000494:pharmacology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013116:Spinal Cord / Q000187:drug effects / Q000378:metabolism*
D002364:Caseins; D015853:Cysteine Proteinase Inhibitors; D003598:Cytoskeletal Proteins; D004151:Dipeptides; D004791:Enzyme Inhibitors; D009185:Myelin Proteins; D018696:Neuroprotective Agents; C026018:azocasein; C087990:AK 275; C090138:AK 295; D002154:Calpain